Objective:To determine whether 5-azacytidine(5-aza) can induce bone marrow mesenchymal stem cells (MSCs) differentiation into cardiomyocytes and whether variant concentration and incubation time of 5-aza have any effect on the differentiation of MSCs.Methods:MSCs were isolated from adult Wistar rats’ femoral and tibial bones and were purified by consecutive passages.The purified MSCs were then treated with 5-aza for 24h in variant concentretions (groupC1-C4:3

5

10

20μmol/L respectively)or in the same concentration(10μmol/L5-aza) but differs from incubation time (groupT1-T5:incubated for 6h

12h

24h

48h

72h respectively).The induced MSCs differentiation was observed by phase-contrast microscope

immunohistochemistry staining and RT-PCR

and the rates of the differentiated MSCs were calculated in each group.Results:Treated with 5-aza

the shape and the aligment pattern of the cells changed gradually

some induced MSCs were stained positive with Desmin and α-Sarcomeric Actin antibody.The rates were 29%

31%

36%

34.5%( P >0.05) and 26%

28.5%

30%

32% ( P >0.05) respectively in groupC1-C4

while in groups T1-T5

the rates were 21%

28%

36%

40%

37.5% ( P <0.01) and 16%

25%

30%

34.5%

33% respectively( P <0.01).The GATA-binding protein 4(GATA4)mRNA expression was detected at the time point 2w

4w and 8w after 5-aza treatment while cardiac troponin T (cTnT) mRNA expressed at the forth week and significantly increased at the eighth week.Conclusion:Induced by appropriate concentration of 5-aza

MSCs of the adult rat can differentiate into cardiomyocyte-like cells.With the increasing 5-aza concentration from 3 μmol/L to 20 μmol/L

the differentiation rates of MSCs were affected slightly( P >0.05).However

the differentiation rate of MSCs increased probably with the incubation time prolonged.During the procedure of MSCs differentiation

the expression pattern of some genes such as GATA4 and cTNT may make some difference.